Proposed work for the coming year includes the investigation of modern approaches for the purification of the transcriptional enzymes from eukaryotic organisms. In addition, the enzymes will be characterized in detail concerning their physical, chemical and enzymatic properties. Protein modifications of these transcriptional components will be monitored during physiological transitions by high resolution techniques, when marked changes in the in vivo rates of RNA synthesis are observed. In vitro reconstituted transcriptional systems will be developed using purified components in order to define what components are necessary for faithful transcription. Highly refined systems will be complemented with other protein fractions in order to ascertain if control 'factors' are required in addition to purified RNA polymerases and templates for faithful transcription. Reconsitution experiments using RNA polymerases which may be modified in vivo when RNA synthesis is activated will also be used in these systems in order to determine the extent to which RNA synthesis may be controlled at the level of the RNA polymerase molecule itself.